Molecular - PCR

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Molecular Pathology

Polymerase Chain Reaction

History

Author: Rodney E. Shackelford, DO, Ph.D. (see Authors page)

Revised: 22 September 2012, last major update January 2010

Pre-PCR history

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● Prior to PCR technology, obtaining multiple copies of a specific DNA sequence was commonly done but often laborious

● Most protocols involved:

- (a) isolating many copies of the sequence(s) desired,

- (b) cloning the DNA into a viral or bacterial plasmid vector

- (c) transfecting, selecting, and growing the bacteria carrying the DNA and vector, and

- (d) re-isolating the desired sequence(s) from bacterial cultures by purifying and cutting the plasmids

● Limitations:

- (a) procedure could take several weeks

- (b) often difficult to get pure DNA/gene sequences from the complex mixtures typically used to obtain DNA samples

Initial discovery of PCR

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● Conceived in 1983 by Kary Mullis (Wikipedia), working at Cetus Corporation as a chemist

● Initial idea was to use a pair of primers to bracket the desired DNA sequence and to copy it repeatedly using DNA polymerase

● Mullis received a $10,000 bonus from Cetus for the invention; Cetus later sold the patent to Roche for $300 million; Mullis may have received additional money for testifying on behalf of Cetus in a patent lawsuit

Later PCR related work

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● Cetus initially used PCR to detect the hemoglobin sickle cell point mutation

● Mullis thought of using DNA polymerase from Thermophilus aquaticus (Taq) (Wikipedia), which was heat resistant; this eliminated the need to add enzyme after every cycle of thermal denaturation of the DNA, making the technique more affordable and subject to automation

● Mullis won the Nobel Prize in Chemistry in 1993 for this work

End of Molecular Pathology > Polymerase Chain Reaction > History

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